PROJECT SUMMARY Itch is defined as an unpleasant sensation that triggers the desire to scratch. During acute itch, scratching only eventually becomes depression. itch/scratching not helps to get rid of pruritogens but also triggers strong mechanical sensation or mechanical pain) that suppresses the itch sensation and stops f urther scratching. During chronic itch, however, scratching uncontrollable, which leads to excessive scratching, skin lesions, anxiety, sleep deprivation, and At present, how scratching information is sensed in the skin and processed in the brain to suppress is largely unknown. ( We find that lateral habenula (LHb) neurons are bilaterally activated after acute pruritogen administration to one side of the mouse cheek, indicating that LHb neurons may respond to chemical itch sensation and/or scratching-induced mechanosensation. Interestingly, LHb neuronal activation is significantly reduced in the contralateral side when mice wear collars to prevent scratching, which suggests that some scratching information is preferentially transmitted to the contralateral LHb. Moreover, re-activation of either bilateral or contralateral itch/scratching-evoked LHb neurons suppresses scratching. Thus, we raise a novel hypothesis that the LHb integrates both scratching-induced mechanosensory and chemical itch sensory inputs and suppress scratching in a lateralized manner and that cutaneous high-threshold mechanoreceptors are required to transmit scratching information to the LHb. We will use a combination of cutting-edge techniques to test this hypothesis. In Aim 1, we will thoroughly examine molecular, circuit, and physiological properties of itch/scratching-activated LHb neurons on the contralateral vs ipsilateral side, with or without collar. We will also record population calcium signal changes of LHb neurons in vivo in response to scratching. In Aim 2, we will test the functional sufficiency and requirement of LHb neurons in suppressing scratching evoked by acute and chronic itch using chemogenetic and optogenetic manipulations as well as LHb neuronal ablation. Finally, in Aim 3, we will ablate C and/or Aδ high threshold mechanoreceptors (HTMRs), which mediate strong mechanical or mechanical pain sensation, to test their functional requirements in transmitting scratching information to the LHb and suppressing scratching. Taken together, our anticipated results will reveal novel neural circuits in mediating the sensation of scratching and controlling scratch, an exciting area that is largely unexplored but highly relevant for alleviating chronic itch.